Construction method of meat analogue based on hypha solid state fermentation

Abstract

The invention relates to the field of fungus solid-state fermentation, and provides a mycelium-based solid-state fermentation meat analogue construction method which comprises the following steps: (1) strain preparation; (2) preparing a solid culture medium; (3) mycelium inoculation; (4) constructing a hypha growth bracket; and (5) mycelium fermentation. According to the method, the structure of meat is simulated in the form of filling the 3D printing bracket with the mycelia through solid-state fermentation growth, so that the meat analogue with the texture and taste similar to those of real meat is created.

Description

Technical Field

[0001] The invention relates to the field of fungal solid-state fermentation, and in particular to a method for constructing meat analogs based on mycelium solid-state fermentation. Background Art

[0002] Meat food is the main source of protein for humans. With the growth of population and the improvement of living standards, the consumption of meat continues to grow. The current meat production and consumption model has caused tremendous pressure on land resources, environment and animal welfare. Therefore, in order to solve these problems, finding sustainable and environmentally friendly meat substitutes has become a hot topic in the global food industry.

[0003] Edible fungus mycelium has a filamentous fiber structure similar to muscle fiber and is rich in high-quality protein, polysaccharides and other nutrients. These bioactive elements can not only improve taste and flavor, but also have potential functional properties. In view of the advantages of mycelium, companies and researchers are exploring innovative methods to incorporate mycelium into meat substitutes. Patent document CN114258978A discloses an artificial meat based on mycelium protein and a preparation method thereof. The method mainly obtains a large amount of mycelium by liquid deep fermentation of edible fungus seeds, mixes the mycelium with gluten, egg white, flavors, etc., and molds after enzyme treatment. Patent document CN113412932A discloses a processing method and application of edible mycelium protein. The method mixes liquid fermented Fusarium mycelium with extruded structured plant protein after denucleation and dehydration, and then adds non-animal fat, pigment, seasoning, food adhesive and edible flavor to obtain a mixture, and the mixture is subjected to quick freezing and steaming and baking to obtain a flavor-adjusted meat substitute. Although the above methods can produce artificial meat rich in mycelium, they are all formed through molds, and their texture characteristics are still somewhat different from those of real meat.

[0004] Therefore, a new method for constructing mycelium meat is needed to meet the demand. Summary of the invention

[0005] The present invention provides a method for constructing meat analogs based on mycelium solid-state fermentation, which constructs a plant protein-based mycelium growth scaffold through 3D printing, and the mycelium fermentation growth and filling obtains mycelium solid-state fermentation meat analogs with ideal texture, taste and flavor.

[0006] A method for constructing meat analogs based on mycelium solid-state fermentation according to the present invention is characterized in that it comprises the following steps:

[0007] (1) Preparation of strains: activating strains using potato dextrose agar medium, and then performing potato dextrose water liquid amplification culture to obtain mycelium; homogenizing and dispersing the mycelium, and then centrifuging to remove the medium components, and then washing with sterile physiological saline, and then centrifuging and dehydrating to collect the mycelium;

[0008] (2) Preparation of solid culture medium: mixing plant protein, carbon source, water, food coloring, flavoring agent and texture improver, heating and then cooling to room temperature to obtain mycelium solid culture medium;

[0009] (3) Mycelium inoculation: The mycelium, TG enzyme and solid culture medium are mixed evenly to prepare a solid culture medium containing mycelium, and the culture medium is loaded into a sterile 3D printing cylinder and refrigerated at 4-8°C for later use;

[0010] (4) Construction of mycelium growth scaffold: construct a culture scaffold model using 3D modeling software; construct a mycelium growth scaffold using the mycelium-containing solid culture medium prepared in step (3) by 3D printing;

[0011] (5) Mycelium fermentation: The constructed mycelium growth scaffold is placed in an incubator for fermentation to obtain mycelium solid-state fermentation meat analogs.

[0012] Preferably, in step (1), the strain is one or more of Pleurotus eryngii, Ganoderma lucidum, Lentinula edodes, Morchella oleracea, Pleurotus ostreatus, and Alpinia cerevisiae.

[0013] Preferably, in step (1), the specific steps are as follows:

[0014] a) inoculating the strain on potato dextrose agar medium, the potato dextrose agar medium formula: 8g potato powder, 2g peptone, 5g maltose, 15g glucose, 1g potassium dihydrogen phosphate, 0.5g magnesium sulfate heptahydrate, 0.05g vitamin B1, 15g agar, 1000ml distilled water, sterilizing at 121°C for 15 minutes; culturing at 24-28°C for 10-15 days, and activating to obtain the primary strain;

[0015] b) using an agar puncher to take 3-5 agar blocks with a diameter of 5-8 mm from the part where the mycelium grows vigorously, and inoculate them into a shaking bottle containing 300 ml of liquid culture medium, the liquid culture medium formula: 8 g potato extract powder, 2 g peptone, 5 g maltose, 15 g glucose, 1 g potassium dihydrogen phosphate, 0.5 g magnesium sulfate heptahydrate, 0.05 g vitamin B1, 1000 ml of distilled water, sterilize at 121° C. for 15 minutes; culture at 24-28° C. and 120-150 rpm on a shaker for 7-10 days; use a sterile homogenizer to homogenize and disperse the mycelium, with a tapping speed of 6-10 times / second and homogenization for 2-4 minutes to obtain a secondary mycelium dispersion;

[0016] c) inoculating the secondary seed mycelium dispersion into a shake flask filled with liquid culture medium at an inoculation rate of 1-3%, culturing at 24-28° C. and 120-150 rpm for 4-6 days to obtain mycelium with a diameter of 3-10 mm, i.e., the tertiary seed;

[0017] d) homogenizing and dispersing the third-grade mycelium with a sterile homogenizer at a tapping speed of 6-10 times / second for 2-4 minutes to obtain a mycelium dispersion; centrifuging the mycelium dispersion at 7000-8000 g for 10 minutes, centrifuging twice to remove the culture medium components; washing the mycelium with sterile physiological saline and centrifuging at 7000-8000 g for 25 minutes, centrifuging twice to remove moisture, and finally collecting the mycelium and refrigerating at 4-8°C for later use.

[0018] Preferably, in step (2), the flavoring agent comprises one or more of pork flavor, beef flavor, and chicken flavor; and the food coloring agent comprises one or more of monascus red, betalain, and capsicum red.

[0019] Preferably, in step (2), the plant protein is one or more of soy protein isolate, wheat gluten, pea protein isolate, corn protein, chickpea protein, and mung bean protein, and the protein content is 85-95%; the carbon source is one or more of soy dietary fiber, oat dietary fiber, starch, D-xylose, and L-arabinose; and the texture modifier is one or more of xanthan gum, curdlan, guar gum, locust bean gum, and konjac glucomannan.

[0020] Preferably, the vegetable protein, carbon source, texture improver, flavoring agent and edible coloring are sterilized by high pressure sterilization at 121° C. for 15 minutes or electron beam irradiation sterilization with an irradiation dose of 10-15 kGy.

[0021] Preferably, in step (2), 18-22wt% of vegetable protein, 4-10wt% of carbon source, 0.1-1wt% of flavoring agent, 0.1-0.5wt% of food coloring, 0.1-2.0wt% of texture improver and 64.5-77.7wt% of sterile water are mixed and stirred at a speed of 400-600rpm for 40-50 minutes using an agitator; the uniformly stirred culture medium is heated at 80-90°C for 30-50 minutes and then cooled to room temperature.

[0022] Preferably, in step (3), TG enzyme sterilized by filtration through a 0.22 μm microporous filter membrane is added to the culture medium at a dosage of 1-5 U per gram of protein at 4-8°C, and then 5-10 wt% of mycelium is added, stirred evenly with a hand-held blender, loaded into a sterile 3D printing barrel, and refrigerated at 4-8°C for use, with a storage time not exceeding 4 hours.

[0023] Preferably, in step (4), the 3D printing parameters are: nozzle diameter is 0.8-1.2 mm, layer height is 70-85% of the nozzle diameter, printing internal filling rate is 30-60%, and printing rate is 15-20 mm / s.

[0024] Preferably, in step (5), the culture conditions in the incubator are: temperature 24-28°C, humidity 85-95%, and culture time 10-15 days.

[0025] The beneficial effects of the present invention are as follows:

[0026] The structure of meat is simulated by filling the 3D printed scaffold with mycelium solid-state fermentation, creating a meat analog with a texture and taste similar to real meat. Mycelium is rich in nutrients such as protein and cellulose, is a good source of protein, and provides dietary fiber, which is beneficial to human health. The mycelium solid-state fermentation process reduces the undesirable characteristics of plant protein, improves flavor and enhances nutritional properties. 3D printing technology combined with mycelium cultivation can customize meat substitutes of different shapes, sizes and nutritional content according to consumer needs. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] Figure 1 This is a flow chart of a method for constructing a meat analog based on mycelium solid-state fermentation in Example 1;

[0028] Figure 2 This is a schematic diagram of the primary species in Example 2;

[0029] Figure 3 This is a schematic diagram of the secondary species in Example 2;

[0030] Figure 4 This is a schematic diagram of the three-level species in Example 2;

[0031] Figure 5 This is a schematic diagram of mycelium in Example 2;

[0032] Figure 6 This is a schematic diagram of the 3D printed Pleurotus eryngii culture medium support in Example 2;

[0033] Figure 7 This is a schematic diagram of the solid-state fermentation of meat analogs by mycelium in Example 2. DETAILED DESCRIPTION

[0034] In order to further understand the content of the present invention, the present invention is described in detail in conjunction with the accompanying drawings and embodiments. It should be understood that the embodiments are only for explaining the present invention and are not intended to limit it.

[0035] Example 1

[0036] like Figure 1As shown, this embodiment provides a method for constructing meat analogs based on mycelium solid-state fermentation, characterized in that it includes the following steps:

[0037] (1) Preparation of strains: The strains were activated using potato dextrose agar medium, and then the activated strains were cultured in potato dextrose water to obtain mycelium; the mycelium was homogenized and dispersed, and then centrifuged to remove the medium components, and then washed with sterile saline, and the mycelium was collected after centrifugal dehydration.

[0038] In step (1), the strains are one or more of Pleurotus eryngii, Ganoderma lucidum, Lentinus edodes, Morchella oleraceus, Pleurotus ostreatus, and Alpinia fasciata.

[0039] In step (1), the specific steps are as follows:

[0040] a) inoculating the strain on potato dextrose agar medium, the potato dextrose agar medium formula: 8g potato powder, 2g peptone, 5g maltose, 15g glucose, 1g potassium dihydrogen phosphate, 0.5g magnesium sulfate heptahydrate, 0.05g vitamin B1, 15g agar, 1000ml distilled water, sterilizing at 121°C for 15 minutes; culturing at 24-28°C for 10-15 days, and activating to obtain the primary strain;

[0041] b) using an agar puncher to take 3-5 agar blocks with a diameter of 5-8 mm from the part where the mycelium grows vigorously, and inoculate them into a shaking bottle containing 300 ml of liquid culture medium, the liquid culture medium formula: 8 g potato extract powder, 2 g peptone, 5 g maltose, 15 g glucose, 1 g potassium dihydrogen phosphate, 0.5 g magnesium sulfate heptahydrate, 0.05 g vitamin B1, 1000 ml of distilled water, sterilize at 121° C. for 15 minutes; culture at 24-28° C. and 120-150 rpm on a shaker for 7-10 days; use a sterile homogenizer to homogenize and disperse the mycelium, with a tapping speed of 6-10 times / second and homogenization for 2-4 minutes to obtain a secondary mycelium dispersion;

[0042] c) inoculating the secondary seed mycelium dispersion into a shake flask filled with liquid culture medium at an inoculation rate of 1-3%, culturing at 24-28° C. and 120-150 rpm for 4-6 days to obtain mycelium with a diameter of 3-10 mm, i.e., the tertiary seed;

[0043] d) homogenizing and dispersing the third-grade mycelium with a sterile homogenizer at a tapping speed of 6-10 times / second for 2-4 minutes to obtain a mycelium dispersion; centrifuging the mycelium dispersion at 7000-8000 g for 10 minutes, centrifuging twice to remove the culture medium components; washing the mycelium with sterile physiological saline and centrifuging at 7000-8000 g for 25 minutes, centrifuging twice to remove moisture, and finally collecting the mycelium and refrigerating at 4-8°C for later use.

[0044] (2) Preparation of solid culture medium: Mix and stir the plant protein, carbon source, water, food coloring, flavoring agent and texture improver, heat and then cool to room temperature to obtain a mycelium solid culture medium.

[0045] In step (2), the flavoring agent includes one or more of pork flavor, beef flavor, and chicken flavor; the food coloring agent includes one or more of monascus red, betalain, and capsicum red.

[0046] In step (2), the plant protein is one or more of soy protein isolate, wheat gluten, pea protein isolate, corn protein, chickpea protein, and mung bean protein, and the protein content is 85-95%; the carbon source is one or more of soy dietary fiber, oat dietary fiber, starch, D-xylose, and L-arabinose; and the texture modifier is one or more of xanthan gum, curdlan, guar gum, locust bean gum, and konjac glucomannan.

[0047] The vegetable protein, carbon source, texture improver, flavoring agent and edible pigment are sterilized by high pressure sterilization at 121° C. for 15 minutes or electron beam irradiation sterilization with an irradiation dose of 10-15 kGy.

[0048] In step (2), 18-22wt% of vegetable protein, 4-10wt% of carbon source, 0.1-1wt% of flavoring agent, 0.1-0.5wt% of food coloring, 0.1-2.0wt% of texture modifier and 64.5-77.7wt% of sterile water are mixed, and stirred at a speed of 400-600rpm for 40-50 minutes using a stirrer; the uniformly stirred culture medium is heated at 80-90°C for 30-50 minutes, and then cooled to room temperature.

[0049] (3) Mycelium inoculation: The mycelium, TG enzyme and solid culture medium are mixed evenly to prepare a solid culture medium containing mycelium. The culture medium is loaded into a sterile 3D printing barrel and refrigerated at 4-8°C for later use.

[0050] In step (3), TG enzyme sterilized by filtration through a 0.22 μm microporous filter membrane is added to the culture medium at a dosage of 1-5 U per gram of protein at 4-8°C, and then 5-10 wt% of mycelium is added. After being stirred evenly with a hand-held blender, it is loaded into a sterile 3D printing barrel and refrigerated at 4-8°C for use. The storage time shall not exceed 4 hours.

[0051] (4) Construction of mycelium growth scaffold: Use 3D modeling software to construct a culture scaffold model; construct a mycelium growth scaffold using the mycelium-containing solid culture medium prepared in step (3) through 3D printing.

[0052] In step (4), the 3D printing parameters are: nozzle diameter is 0.8-1.2 mm, layer height is 70-85% of the nozzle diameter, printing internal filling rate is 30-60%, and printing speed is 15-20 mm / s.

[0053] (5) Mycelium fermentation: The constructed mycelium growth scaffold is placed in an incubator for fermentation to obtain mycelium solid-state fermentation meat analogs.

[0054] In step (5), the culture conditions of the incubator are: temperature 24-28° C., humidity 85-95%, and culture time 10-15 days.

[0055] The structure of meat is simulated by filling the 3D printed scaffold with mycelium solid-state fermentation, creating a meat analog with a texture and taste similar to real meat. Mycelium is rich in nutrients such as protein and cellulose, is a good source of protein, and provides dietary fiber, which is beneficial to human health. The mycelium solid-state fermentation process reduces the undesirable characteristics of plant protein, improves flavor and enhances nutritional properties. 3D printing technology combined with mycelium cultivation can customize meat substitutes of different shapes, sizes and nutritional content according to consumer needs.

[0056] Example 2

[0057] In this embodiment, the strain is Pleurotus eryngii, and the strain mother is inoculated on an agar plate medium and cultured at 25° C. for 12 days to activate and obtain a primary strain. Figure 2 shown.

[0058] Use an agar puncher to take 5 agar blocks with a diameter of 8 mm from the vigorous mycelial growth area, inoculate them into a shake flask containing 300 ml of potato dextrose water liquid culture medium, and culture at 25°C and 150 rpm for 10 days. Figure 3 The mycelium was homogenized and dispersed using a sterile homogenizer, with a tapping speed of 6 times / second and homogenization for 3 minutes to obtain a secondary mycelium dispersion.

[0059] The secondary seed mycelium dispersion was inoculated into a shake flask filled with potato glucose water liquid culture medium at an inoculum of 1.5%, and cultured at 25°C and 150 rpm for 6 days to obtain mycelium with a diameter of 3-10 mm, i.e., the third seed. Figure 4 shown.

[0060] Use a sterile homogenizer to homogenize and disperse the third-level seeds at a tapping speed of 8 times / second for 3 minutes to obtain a mycelium dispersion. Centrifuge the mycelium dispersion at 8000g for 10 minutes, centrifuge twice to remove the culture medium components; wash the mycelium with sterile saline and centrifuge at 8000g for 25 minutes, centrifuge twice to remove water, and finally collect the mycelium and refrigerate at 4-8℃ for later use. Figure 5 shown.

[0061] Beef flavor was sterilized by electron beam irradiation with a dose of 10 kGy. Soy protein isolate, tapioca starch and curdlan were sterilized by electron beam irradiation with a dose of 15 kGy.

[0062] 18 wt% of soy protein isolate, 5 wt% of tapioca starch, 0.5 wt% of curdlan, 0.1 wt% of beef flavor, and 76.4 wt% of sterile water were mixed, and stirred at 500 rpm using a stirrer for 40 minutes; the uniformly stirred culture medium was heated at 80° C. for 30 minutes, and then cooled to room temperature.

[0063] At 4-8°C, TG enzyme sterilized by filtration through a 0.22 μm microporous filter membrane was added to the culture medium at a dosage of 2 U per gram of protein, and then 6 wt% of mycelium was added. After stirring evenly with a hand-held blender, it was loaded into a sterile printing barrel and refrigerated at 4-8°C for later use.

[0064] The mycelium growth scaffold was constructed by 3D printing, such as Figure 6 As shown, the 3D printing parameters are: nozzle diameter is 1mm, layer height is 0.84mm; printing internal filling rate is 40%, and printing speed is 15mm / s.

[0065] The constructed mycelium growth scaffold is placed in an incubator for fermentation to obtain mycelium solid-state fermentation meat analogs, such as Figure 7 The culture conditions of the incubator are: temperature 24-28°C, humidity 85-95%, and culture time 15 days.

[0066] Example 3

[0067] In this embodiment, the mushroom strain is Pleurotus ostreatus, and the mother strain is inoculated on an agar plate medium, cultured at 25° C. for 12 days, and activated to obtain a primary strain.

[0068] Use an agar puncher to take 5 agar blocks with a diameter of 8 mm from the area where the mycelium is growing vigorously, inoculate them into a shake flask containing 300 ml of liquid culture medium, and culture them at 25°C and 150 rpm on a shaker for 10 days. Use a sterile homogenizer to homogenize and disperse the mycelium at a tapping speed of 6 times / second and homogenize for 3 minutes to obtain a secondary mycelium dispersion.

[0069] The secondary seed mycelium dispersion was inoculated into a shaking flask filled with liquid culture medium at an inoculation rate of 1.5%, and cultured at 25° C. and 150 rpm for 6 days to obtain mycelium with a diameter of 3-10 mm, namely the tertiary seed.

[0070] Use a sterile homogenizer to homogenize and disperse the third-grade mycelium, beat at a speed of 8 times / second, and homogenize for 3 minutes to obtain a mycelium dispersion. Centrifuge the mycelium dispersion at 8000g for 10 minutes, centrifuge twice, and remove the culture medium components; wash the mycelium with sterile physiological saline and centrifuge at 8000g for 25 minutes, centrifuge twice, remove water, and finally collect the mycelium and refrigerate at 4-8℃ for later use.

[0071] Chicken flavor and food coloring were sterilized by electron beam irradiation with a dose of 10 kGy. Pea protein, tapioca starch and xanthan gum were sterilized by electron beam irradiation with a dose of 15 kGy.

[0072] 21wt% of pea protein, 6wt% of cassava starch, 0.1% of xanthan gum, 0.1wt% of chicken meat flavor, 0.1% of monascus red, and 72.7wt% of sterile water were mixed, and stirred at 500rpm for 40 minutes using a stirrer; the evenly stirred culture medium was heated at 80°C for 30 minutes, and then cooled to room temperature.

[0073] At 4-8°C, TG enzyme sterilized by filtration through a 0.22 μm microporous filter membrane was added to the culture medium at a dosage of 2 U per gram of protein, and then 6 wt% of mycelium was added. After stirring evenly with a hand-held blender, the mixture was loaded into a sterile printing cylinder and refrigerated at 4-8°C for later use.

[0074] The 3D printing parameters are: nozzle diameter is 1mm, layer height is 0.84mm; printing internal filling rate is 40%, and printing speed is 15mm / s.

[0075] The constructed mycelium growth support is placed in an incubator for fermentation to obtain mycelium solid-state fermented meat analogs. The incubator culture conditions are: temperature 24-28°C, humidity 85-95%, and culture time 15 days.

[0076] Example 4

[0077] In this example, the hardness of the mycelium fermented meat analogs prepared in Examples 2 and 3 was compared with that of chicken and beef, as shown in Table 1:

[0078] Table 1 Hardness of mycelium fermented meat analogs

[0079] Hardness(N) Example 2 29.72 Example 3 19.73 chicken 22.52 beef 32.39

[0080] As can be seen from Table 1, the mycelium fermented meat analogs prepared in Examples 2 and 3 have similar hardness to chicken and beef, and can be used as good meat substitutes.

[0081] The present invention and its embodiments are described schematically above, and the description is not restrictive. The drawings show only one embodiment of the present invention, and the actual structure is not limited thereto. Therefore, if a person skilled in the art is inspired by it and designs a structural method and an embodiment similar to the technical solution without creativity without departing from the purpose of the invention, they shall all fall within the protection scope of the present invention.

Claims

1. A method for constructing meat analogs based on mycelium solid-state fermentation, characterized in that: The following steps are involved: (1) Preparation of strains: activating strains using potato dextrose agar medium, and then performing potato dextrose water liquid amplification culture to obtain mycelium; homogenizing and dispersing the mycelium, and then centrifuging to remove the medium components, and then washing with sterile physiological saline, and then centrifuging and dehydrating to collect the mycelium; (2) Preparation of solid culture medium: mixing plant protein, carbon source, water, food coloring, flavoring agent and texture improver, heating and then cooling to room temperature to obtain mycelium solid culture medium; (3) Mycelium inoculation: The mycelium, TG enzyme and solid culture medium are mixed evenly to prepare a solid culture medium containing mycelium, and the culture medium is loaded into a sterile 3D printing cylinder and refrigerated at 4-8°C for later use; (4) Construction of mycelium growth scaffold: construct a culture scaffold model using 3D modeling software; construct a mycelium growth scaffold using the mycelium-containing solid culture medium prepared in step (3) by 3D printing; (5) Mycelium fermentation: The constructed mycelium growth scaffold is placed in an incubator for fermentation to obtain mycelium solid-state fermentation meat analogs.

2. The method for constructing meat analogs based on mycelium solid-state fermentation according to claim 1, characterized in that: In step (1), the strains are one or more of Pleurotus eryngii, Ganoderma lucidum, Lentinus edodes, Morchella oleraceus, Pleurotus ostreatus, and Alpinia fasciata.

3. The method for constructing meat analogs based on mycelium solid-state fermentation according to claim 2, characterized in that: In step (1), the specific steps are as follows: a) inoculating the strain on potato dextrose agar medium, the potato dextrose agar medium formula: 8g potato powder, 2g peptone, 5g maltose, 15g glucose, 1g potassium dihydrogen phosphate, 0.5g magnesium sulfate heptahydrate, 0.05g vitamin B1, 15g agar, 1000ml distilled water, sterilizing at 121°C for 15 minutes; culturing at 24-28°C for 10-15 days, and activating to obtain the primary strain; b) using an agar puncher to take 3-5 agar blocks with a diameter of 5-8 mm from the part where the mycelium grows vigorously, and inoculate them into a shaking bottle containing 300 ml of liquid culture medium, the liquid culture medium formula: 8 g potato extract powder, 2 g peptone, 5 g maltose, 15 g glucose, 1 g potassium dihydrogen phosphate, 0.5 g magnesium sulfate heptahydrate, 0.05 g vitamin B1, 1000 ml of distilled water, sterilize at 121° C. for 15 minutes; culture at 24-28° C. and 120-150 rpm on a shaker for 7-10 days; use a sterile homogenizer to homogenize and disperse the mycelium, with a tapping speed of 6-10 times / second and homogenization for 2-4 minutes to obtain a secondary mycelium dispersion; c) inoculating the secondary seed mycelium dispersion into a shake flask filled with liquid culture medium at an inoculation rate of 1-3%, culturing at 24-28° C. and 120-150 rpm for 4-6 days to obtain mycelium with a diameter of 3-10 mm, i.e., the tertiary seed; d) homogenizing and dispersing the third-grade mycelium with a sterile homogenizer at a tapping speed of 6-10 times / second for 2-4 minutes to obtain a mycelium dispersion; centrifuging the mycelium dispersion at 7000-8000 g for 10 minutes, centrifuging twice to remove the culture medium components; washing the mycelium with sterile physiological saline and centrifuging at 7000-8000 g for 25 minutes, centrifuging twice to remove moisture, and finally collecting the mycelium and refrigerating at 4-8°C for later use.

4. The method for constructing meat analogs based on mycelium solid-state fermentation according to claim 3, characterized in that: In step (2), the flavoring agent includes one or more of pork flavor, beef flavor, and chicken flavor; the food coloring agent includes one or more of monascus red, betalain, and capsicum red.

5. The method for constructing meat analogs based on mycelium solid-state fermentation according to claim 4, characterized in that: In step (2), the plant protein is one or more of soy protein isolate, wheat gluten, pea protein isolate, corn protein, chickpea protein, and mung bean protein, and the protein content is 85-95%; the carbon source is one or more of soy dietary fiber, oat dietary fiber, starch, D-xylose, and L-arabinose; and the texture modifier is one or more of xanthan gum, curdlan, guar gum, locust bean gum, and konjac glucomannan.

6. The method for constructing meat analogs based on mycelium solid-state fermentation according to claim 5, characterized in that: The vegetable protein, carbon source, texture improver, flavoring agent and edible pigment are sterilized by high pressure sterilization at 121° C. for 15 minutes or electron beam irradiation sterilization with an irradiation dose of 10-15 kGy.

7. The method for constructing meat analogs based on mycelium solid-state fermentation according to claim 6, characterized in that: In step (2), 18-22wt% of vegetable protein, 4-10wt% of carbon source, 0.1-1wt% of flavoring agent, 0.1-0.5wt% of food coloring, 0.1-2.0wt% of texture modifier and 64.5-77.7wt% of sterile water are mixed, and stirred at a speed of 400-600rpm for 40-50 minutes using a stirrer; the uniformly stirred culture medium is heated at 80-90°C for 30-50 minutes, and then cooled to room temperature.

8. The method for constructing meat analogs based on mycelium solid-state fermentation according to claim 7, characterized in that: In step (3), TG enzyme sterilized by filtration through a 0.22 μm microporous filter membrane is added to the culture medium at a dosage of 1-5 U per gram of protein at 4-8°C, and then 5-10 wt% of mycelium is added. After being stirred evenly with a hand-held blender, it is loaded into a sterile 3D printing barrel and refrigerated at 4-8°C for use. The storage time shall not exceed 4 hours.

9. The method for constructing meat analogs based on mycelium solid-state fermentation according to claim 8, characterized in that: In step (4), the 3D printing parameters are: nozzle diameter is 0.8-1.2 mm, layer height is 70-85% of the nozzle diameter, printing internal filling rate is 30-60%, and printing speed is 15-20 mm / s.

10. The method for constructing meat analogs based on mycelium solid-state fermentation according to claim 9, characterized in that: In step (5), the culture conditions of the incubator are: temperature 24-28° C., humidity 85-95%, and culture time 10-15 days.

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